Mohammad Ghader Bayazidi
1, Reza Rahbarghazi
2,3 
, Aysa Rezabakhsh
4 , Jafar Rezaie
5 , Mehdi Hassanpour
2, Mahdi Ahmadi
2,1* 1 Department of Physiology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran
2 Stem Cell Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
3 Department of Applied Cell Sciences, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran
4 Cardiovascular Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
5 Solid Tumor Research Center, Research Institute for Cellular and Molecular Medicine, Urmia University of Medical Sciences, Urmia, Iran
Abstract
Introduction: The current experiment aimed to address the impact of type 2 diabetes mellitus on autophagy status in the rat pulmonary tissue.
Methods: In this study, 20 male Wistar rats were randomly allocated into two groups as follows: control and diabetic groups. To induce type 2 diabetes mellitus, rats received a combination of streptozotocin (STZ) and a high-fat diet. After confirmation of diabetic condition, rats were maintained for 8 weeks and euthanized for further analyses. The pathological changes were assessed using H&E staining. We also measured transforming growth factor-β (TGF-β), bronchoalveolar lavage fluid (BALF), and tumor necrosis factor-α (TNF-α) in the lungs using ELISA and real-time PCR analyses, respectively. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were monitored in diabetic lungs to assess oxidative status. We also measured the expression of becline-1, LC3, and P62 to show autophagic response under diabetic conditions. Using immunofluorescence staining, protein levels of LC3 was also monitored.
Results: H&E staining showed pathological changes in diabetic rats coincided with the increase of TNF-α (~1.4-fold) and TGF-β (~1.3-fold) compared to those in the normal rats (P < 0.05). The levels of MDA (5.6 ± 0.4 versus 6.4 ± 0.27 nM/mg protein) were increased while SOD (4.2 ± 0.28 versus 3.8 ± 0.13 U/mL) activity decreased in the diabetic rats (P < 0.05). Real-time polymerase chain reaction (PCR) analysis showed the up-regulation of Becline-1 (~1.35-fold) and LC3 (~2-fold) and down-regulation of P62 (~0.8-fold) (P < 0.05), showing incomplete autophagic flux. We noted the increase of LC3+ cells in diabetic condition compared to that in the control samples.
Conclusion: The prolonged diabetic condition could inhibit the normal activity of autophagy flux, thereby increasing pathological outcomes.